Michel Renouil

In vitro prediction of stop-codon suppression by intravenous gentamicin in patients with cystic fibrosis: a pilot study.

BackgroundCystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) protein, which acts as a chloride channel activated by cyclic AMP (cAMP). The most frequent mutation found in 70% of CF patients is F508del, while premature stop mutations are found in about 10% of patients. In vitro aminoglycoside antibiotics (e.g. gentamicin) suppress nonsense mutations located in CFTR permitting translation to continue to the natural termination codon. Pharmacologic suppression of stop mutations within the CFTR may be of benefit to a significant number of patients. Our pilot study was conducted to determine whether intravenous gentamicin suppresses stop codons in CF patients and whether it has clinical benefits.MethodsA dual gene reporter system was used to determine the gentamicin-induced readthrough level of the most frequent stop mutations within the CFTR in the French population. We investigated readthrough efficiency in response to 10 mg/kg once-daily intravenous gentamicin perfusions in patients with and without stop mutations. Respiratory function, sweat chloride concentration, nasal potential difference (NPD) and CFTR expression in nasal epithelial cells were measured at baseline and after 15 days of treatment.ResultsAfter in vitro gentamicin incubation, the readthrough efficiency for the Y122X mutation was at least five times higher than that for G542X, R1162X, and W1282X. In six of the nine patients with the Y122X mutation, CFTR immunodetection showed protein at the membrane of the nasal epithelial cells and the CFTR-dependent Cl- secretion in NPD measurements increased significantly. Respiratory status also improved in these patients, irrespective of the gentamicin sensitivity of the bacteria present in the sputum. Mean sweat chloride concentration decreased significantly and normalised in two patients. Clinical status, NPD and sweat Cl- values did not change in the Y122X patients with no protein expression, in patients with the other stop mutations investigated in vitro and those without stop mutations.ConclusionSuppression of stop mutations in the CFTR gene with parenteral gentamicin can be predicted in vitro and is associated with clinical benefit and significant modification of the CFTR-mediated Cl- transport in nasal and sweat gland epithelium.

Down-regulation of the Anti-inflammatory Protein Annexin A1 in Cystic Fibrosis Knock-out Mice and Patients

Cystic fibrosis is a fatal human genetic disease caused by mutations in the CFTR gene encoding a cAMP-activated chloride channel. It is characterized by abnormal fluid transport across secretory epithelia and chronic inflammation in lung, pancreas, and intestine. Because cystic fibrosis (CF) pathophysiology cannot be explained solely by dysfunction of cystic fibrosis transmembrane conductance regulator (CFTR), we applied a proteomic approach (bidimensional electrophoresis and mass spectrometry) to search for differentially expressed proteins between mice lacking cftr (cftrtm1Unc, cftr−/−) and controls using colonic crypts from young animals, i.e. prior to the development of intestinal inflammation. By analyzing total proteins separated in the range of pH 6–11, we detected 24 differentially expressed proteins (>2-fold). In this work, we focused on one of these proteins that was absent in two-dimensional gels from cftr−/− mice. This protein spot (molecular mass, 37 kDa; pI 7) was identified by mass spectrometry as annexin A1, an anti-inflammatory protein. Interestingly, annexin A1 was also undetectable in lungs and pancreas of cftr−/− mice, tissues known to express CFTR. Absence of this inhibitory mediator of the host inflammatory response was associated with colonic up-regulation of the proinflammatory cytosolic phospholipase A2. More importantly, annexin A1 was down-regulated in nasal epithelial cells from CF patients bearing homozygous nonsense mutations in the CFTR gene (Y122X, 489delC) and differentially expressed in F508del patients. These results suggest that annexin A1 may be a key protein involved in CF pathogenesis especially in relation to the not well defined field of inflammation in CF. We suggest that decreased expression of annexin A1 contributes to the worsening of the CF phenotype.

Chikungunya virus–associated encephalitis: A cohort study on La Réunion Island, 2005–2009

Objective: To estimate the cumulative incidence rate (CIR) of Chikungunya virus (CHIKV)–associated CNS disease during the La Réunion outbreak, and assess the disease burden and patient outcome after 3 years. Methods: CHIKV-associated CNS disease was characterized retrospectively in a cohort of patients with positive CHIKV reverse transcriptase PCR or anti-CHIKV immunoglobulin M antibodies in the CSF and fulfilling International Encephalitis Consortium criteria for encephalitis or encephalopathy. Neurologic sequelae were assessed after 3 years. Results: Between September 2005 and June 2006, 57 patients were diagnosed with CHIKV-associated CNS disease, including 24 with CHIKV-associated encephalitis, the latter corresponding to a CIR of 8.6 per 100,000 persons. Patients with encephalitis were observed at both extremes of age categories. CIR per 100,000 persons were 187 and 37 in patients below 1 year and over 65 years, respectively, both far superior to those of cumulated causes of encephalitis in the United States in these age categories. The case-fatality rate of CHIKV-associated encephalitis was 16.6% and the proportion of children discharged with persistent disabilities estimated between 30% and 45%. Beyond the neonatal period, the clinical presentation and outcomes were less severe in infants than in adults. Conclusions: In the context of a large outbreak, CHIKV is a significant cause of CNS disease. As with other etiologies, CHIKV-associated encephalitis case distribution by age follows a U-shaped parabolic curve.

Mutation in a primate-conserved retrotransposon reveals a noncoding RNA as a mediator of infantile encephalopathy

The human genome is densely populated with transposons and transposon-like repetitive elements. Although the impact of these transposons and elements on human genome evolution is recognized, the significance of subtle variations in their sequence remains mostly unexplored. Here we report homozygosity mapping of an infantile neurodegenerative disease locus in a genetic isolate. Complete DNA sequencing of the 400-kb linkage locus revealed a point mutation in a primate-specific retrotransposon that was transcribed as part of a unique noncoding RNA, which was expressed in the brain. In vitro knockdown of this RNA increased neuronal apoptosis, consistent with the inappropriate dosage of this RNA in vivo and with the phenotype. Moreover, structural analysis of the sequence revealed a small RNA-like hairpin that was consistent with the putative gain of a functional site when mutated. We show here that a mutation in a unique transposable element-containing RNA is associated with lethal encephalopathy, and we suggest that RNAs that harbor evolutionarily recent repetitive elements may play important roles in human brain development.

Measurement of nasal potential difference in young children with an equivocal sweat test following newborn screening for cystic fibrosis

Background A challenging problem arising from cystic fibrosis (CF) newborn screening is the significant number of infants with hypertrypsinaemia (HIRT) with sweat chloride levels in the intermediate range and only one or no identified CF-causing mutations. Objectives To investigate the diagnostic value for CF of assessing CF transmembrane conductance regulator (CFTR) protein function by measuring nasal potential difference in children with HIRT. Methods A specially designed protocol was used to assess nasal potential difference (NPD) in 23 young children with HIRT (3 months–4 years) with inconclusive neonatal screening. Results were analysed with a composite score including CFTR-dependent sodium and chloride secretion. Results were correlated with genotype after extensive genetic screening and with clinical phenotype at follow-up 3 years later. Results NPD was interpretable for 21 children with HIRT: 13 had NPD composite scores in the CF range. All 13 were finally found to carry two CFTR mutations. At follow-up, nine had developed a chronic pulmonary disease consistent with a CF diagnosis. The sweat test could be repeated in nine children, and six had sweat chloride values ≥60 mmol/l. Of the eight children with normal NPD scores, only two had two CFTR mutations, both wide-spectrum mutations. None had developed a CF-like lung disease at follow-up. The sweat test could be reassessed in five of these eight children and all had sweat chloride values <60 mmol/l. CF diagnosis was ruled out in six of these eight children. Conclusion Evaluation of CFTR function in the nasal epithelium of young children with inconclusive results at CF newborn screening is a useful diagnostic tool for CF.

Déshydratation hypernatrémique et allaitement maternel

Neonatal hypernatremic dehydration due to breast feeding was rarely reported in the French-language literature. Case report. – The authors report hypernatremic dehydration in a 10-day-old exclusively breast-fed infant. The course was favorable. Insufficient breast milk production has been reported for 30 years. Conclusion. – Early discharge from maternity units is frequent nowadays. Breast-fed newborns without a positive weight gain at discharge should be checked for their weight within the first ten days of life.

Two Patients With Imported Acute Neuroschistosomiasis Due to Schistosoma mansoni

We report the case of two brothers who returned from Madagascar presenting all the acute phase symptoms of a primary invasive Schistosoma mansoni infection, together with brain involvement characterized by acute encephalitis. This rarely described issue should be considered in travelers returning from endemic areas with acute neurological symptoms.

Méningites à éosinophiles et angiostrongylose chez le nourrisson à l’Île de la Réunion: à propos de deux cas

Resume Le diagnostic etiologique des meningites a Angiostrongylus cantonensis est difficile. L’ A cantonensis est un nematode dont le cycle parasitaire passe par le rat, l’escargot et la limace. Accidentellement, l’homme infeste constitue une impasse parasitaire. Observation — Les auteurs rapportent deux observations de meningites a A. cantonensis confirmees par la serologie entre 1993 et 1998. Le diagnostic de ces meningites a ete evoque devant une hypereosinophilie dans le sang et dans le liquide cephalorachidien. Le traitement de ces meningites est controverse. Cependant, la corticotherapie et les antihelminthiques (albendazole, thiabendazole, levamisole, praziquantel) restent proposes par certains auteurs. Le pronostic est souvent favorable, mais des formes aigues graves et des sequelles neurologiques ont ete decrites. Conclusion — Devant un syndrome meninge, une hypereosinophilie dans le sang et dans le LCR chez un nourrisson demeurant ou ayant sejourne a l’Ile de la Reunion, il faut evoquer le diagnostic de meningite a A. cantonensis .